This invention was made with government support under a grant from the National Institutes of Health. The government has certain rights to this invention.
This invention relates to methods of making epitopes and nucleic acids embodying the epitopes so made.
It is desireable to be able to probe and dissect the precise sites of antigen-antibody interaction. It is also desireable to find novel ways to detect antibodies and to inhibit specific antibody-antigen interactions. Furthermore, methods are needed that allow one to purify a monospecific antibody from a polyclonal serum without having to first purify the antigen.
It has not heretofore been possible to produce distinct and generally useful epitopes which react with a given antibody except in the case of epitope libraries. These peptide libraries depend upon expression of a random set of epitopes within the context of a larger protein. See, e.g., Scott et al., Science 249, 386 (1990); Cwirla et al., Proc. Natl. Acad. Sci. USA 87, 6378 (1990). This approach is restricted because it offers only proteinaceous ligands and is potentially compromised in contexts other than the fusion protein. In addition, xe2x80x9ccombinatorial peptide librariesxe2x80x9d have been described which apply sequential positional determinations. See Houghten et al., Nature 454, 84 (1991). This procedure requires evaluation of the selected ligand at each step which, in turn, requires an exponential effort to define and select a specific epitope. Further, methods for screening degenerate pools of peptide sequences have been used which are not limited by proteinaceous context but are limited for logistical reasons (e.g., sophisticated synthesis and detection instruments are required). See Fodor et al., Science 251, 767 (1991); Geysen et al., Proc. Natl. Acad. Sci. USA 81, 3998 (1984).
S. Deutscher and J. Keene, Proc. Natl. Acad. Sci. USA 85, 3299 (1988) describe the selection and amplification of a nucleic acid ligand on U1 RNA from a randomized pool of nucleic acids (see also J. Wilusz and J. Keene, J. Biol. Chem. 261, 5467 (1986)). L. Gold and C. Tuerk, Nucleic Acid Ligands, PCT Appln. Publn No. WO 91/19813 (Dec. 26, 1991), describe the xe2x80x9cevolutionxe2x80x9d of nucleic acid ligands and nucleic acid compounds refered to as xe2x80x9cnucleic acid antibodiesxe2x80x9d (see also C. Tuerk and L. Gold, Science 249, 505-510 (1990)). A. Ellington and J. Szostak, Nature 346, 818-822 (1990), describe the binding of RNA molecules to organic dyes. D. Tsai et al., Nucleic Acids Research 19, 4931-4936 (1991), describe the binding of the U1-snRNP-A protein to specific RNA sequences in a degenerate pool of transcripts. D. Bartel et al., Cell 67, 529-563 (1991), describe the binding of the Rev protein of HIV-1 to a nucleic acid pool.
There has not heretofore been described a method by which an antibody can be employed to derive a nonproteinaceous mimetic ligand that binds to the same site on the antibody to which the original antigen bound.
A first aspect of the present invention is a method of generating a nucleic acid molecule which is immunologically cross-reactive with an immunogen, which immunogen is not a nucleic acid (e.g., a peptide). The method comprises combining an antigen binding protein which binds the immunogen (e.g., an antibody, a T cell receptor) with a degenerate pool of nucleic acid species, and then recovering a nucleic acid species bound by said binding protein from the degenerate pool.
A second aspect of the present invention is an isolated nucleic acid which inhibits complex formation between an antigen binding protein and an immunogen, which immunogen is not a nucleic acid. In one embodiment, the nucleic acid inhibits complex formation between a self peptide autoantigen and an antigen binding protein.
The foregoing and other objects and aspects of the present invention are explained in detail in the drawings herein and the specification set forth below.